Superfund Research Program

Protein Characterization

Project Leader: Elizabeth A. Komives
Grant Number: P42ES010337
Funding Period: 2005-2010

Project-Specific Links

Final Progress Reports

Year:   2009 

The protein characterization core has been continuing to work on all three specific aims, and beginning some new developments. In collaboration with the Schroeder lab, they have focused on analyzing the metabolic flux from glutathione biosynthesis to phytochelatins. The quantitative analysis of each metabolite along the way in the presence and absence of heavy metal stress has been performed. The results pinpoint several "control points" in the pathway. Some of the results were recently published:
Sung DY, Kim TH, Komives EA, Mendoza-Cózatl DG, Schroeder JI. ARS5 is a component of the 26S proteasome complex, and negatively regulates thiol biosynthesis and arsenic tolerance in Arabidopsis. Plant J. 2009 Sep;59(5):802-13.

They are continuing to perform analysis of protein complexes that have been isolated by various affinity methods, although now researchers are performing the protein digestion and identification experiments in solution rather than cutting bands out of gels, which has increased the number of proteins they can identify, and can also identify lower-abundance proteins. Using this approach, they were able to identify binding partners of neurexin in collaboration with P. Taylor and D. Comoletti. This protein is involved in autism spectrum disorders, and little is known about its physiological function much less the pathophysiology of autism. Some of the interesting proteins researchers found associated with neurexin were voltage-gated potassium channels and the neuronal pentraxin receptor (among many others).

In new developments, researchers have been establishing protein quantification methods in the lab, focusing on the iTRAQ technology. Several experiments were performed in collaboration with the Schroeder lab to identify changes in phosphorylation upon As treatment, but these results have not yet been published. They have also focused on developing MALDI Tof-tof imaging approaches for identifying glucuronide metabolites since this is of interest to the Tukey lab. A novel MALDI target surface called cation-enhanced nanostructure-initiator mass spectrometry (or NIMS) was developed by G. Suisdak at TSRI and researchers have been collaborating with Dr. Suisdak to implement this surface for glucuronide analysis. They have also established the hardware and software necessary for tissue imaging and expect to be able to merge the two technologies in the coming year.